FIG. 7.
The decrease of free ATP levels in HIF-1α null primary fibroblasts parallels that observed in transformed null cells. Following infection with β-galactosidase or cre adenovirus, primary mEFs were seeded at 2 × 105 to 3 × 105 in DMEM–high-glucose medium supplemented with 25 mM HEPES (pH 7.4). The next day, the cells were left at normoxia or were transferred to hypoxia for 24 h. Cell extracts were prepared and normalized as described in Materials and Methods. At normoxia (solid bars) there was no statistically significant difference in the levels of free ATP production in wild-type or null cells. However, at hypoxia (hatched bars) the decrease in the levels of free ATP produced by null cells was approximately 50%, paralleling the decrease observed in transformed cells as shown in Fig. 6. Therefore, the effect of loss of HIF-1α on cellular metabolism is consistent in both normal and transformed cells.